The development of a neuroprotective/restorative therapy for Parkinson's disease (PD) would be a major therapeutic advance, particularly for our veterans. Glial cell line-derived neurotrophic factor (GDNF) has been the most potent trophic molecule to dopamine neurons affected in PD. GDNF requires focal delivery as it does not cross the blood-brain barrier (BBB). However, clinical trials of GDNF in PD patients have given mixed results. It is challenging to achieve therapeutic levels of GDNF for all or most degenerating neurons due to the relatively large target area in human brain and the poor brain tissue penetration of this molecule. Additionally, current GDNF therapy entails important safety concerns. We recently developed a novel approach - macrophage-mediated GDNF delivery - that seems capable of resolving these problems. This unique approach takes advantage of the well-known macrophage property of homing to degenerating sites in proximity to damaged neurons, capitalizes on our powerful macrophage-specific synthetic promoters (MSP), and implements recent advances in hematopoietic stem cell (HSC) gene therapy. We hypothesize that highly effective CNS delivery of GDNF can be achieved through its expression in macrophages / microglia by ex vivo transduction of HSC-containing bone marrow cells with lentiviral vectors carrying a cassette expressing GDNF driven by our MSP, followed by syngeneic transplantation of these transduced bone marrow cells, and this will greatly ameliorate the pathological changes and neurological defects of PD. Previously, using a sub-acute MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) mouse model of PD in a largely preventive paradigm, we demonstrated that genetically engineered bone marrow HSC-derived macrophages accumulate in diseased sites and macrophage-mediated GDNF delivery dramatically reduces degeneration of dopaminergic neurons of the substantia nigra, as well as their fibers in the striatum, and induces axon regeneration, without any apparent adverse effects. Recently, using the conditional knockout MitoPark mouse as a chronic and progressive model of PD, we showed similar therapeutic results with our hematopoietic stem cell transplantation (HSCT)-based macrophage/microglia-mediated GDNF delivery approach in treatment settings. In addition, the concern that macrophage-infiltration into the brain might be induced by irradiation (as transplant preparative regimen)-caused BBB damage was essentially lifted as the mouse heads were protected in the study. Moreover, PD-like non-motor symptoms were observed in the MitoPark mice and in the preliminary study could be mitigated by our GDNF delivery approach. Very recently, we developed a new gentle and non- cytotoxic HSCT method. Conventional HSCT requires cytotoxic preconditioning such as chemotherapy and may thus not suitable to our veterans with PD. The basis for this new technology is pharmacologic (e.g., G- CSF, AMD3100) stem cell mobilization and clearance of HSC niches in the bone marrow microenvironment, thereby opening the way for high-level engraftment of genetically modified transplanted HSCs. In this grant application, we first propose to further examine the potential therapeutic effects of our GDNF therapy on non- motor symptoms, an unmet need in PD. We then propose to test our macrophage-mediated GDNF delivery approach through the new HSCT method for improvement of neurodegeneration as well as motor and non- motor activities in the mouse model of PD. The combination study is anticipated to lead to a completely innovative neuroprotective/disease-modifying therapy for PD with increased feasibility and therefore establish a solid base for future clinical investigation of the potential benefits to patients of tis PD management.